| Stretch is a type of physiological stimulation in the gastrointestinal smooth muscles. The previous study had showed that mechanical forces could stimulate plenty of signal transduction, including those initiated by or resulting in ion channel activation. Membrane stretch, for example, can potentiate Ca2+, K+ and muscarinic currents. The cytoskeleton is made of microfilament, microtubule and intermediate fiber that form a complex network and provide a basis for simultaneous interactions among multiple cellular structures. Membrane stretch can deform the shape of cell so that it can alter the cytoskeleton structure inevitably. It had been already known that disruption of microfilament inhibited the increase of Ca2+ and KATP induced by the hyposmotic membrane stretch. As the previous study showed, membrane stretch also enhanced muscarinic current whose mechanism had not been clear. Today, it is not clear whether cytoskeleton plays a role in muscarinic current under normal and membrane stretch condition. The aim of this study was to explain whether cytockeleton be involved in membrane stretch-induced increase of muscarinic current. In this study, collagenase Ⅱ (0.1%) was applied to dissociate gastric antral circular smooth muscle cells of the guinea-pig. By the whole-cell configuration of patch-clamp technique, in this study, the relationship between microfilament and muscarinic current under normal and membrane stretch condition had been investigated. Origin3.73 and SigmaPlot 4.01 were used to statistic.Results are as follows:1. By the conventional whole-cell patch-clamp technique, 50 ?mol/L Carbachol (Cch, a muscarinic antagonist) in isosmotic solution elicited an inward current (ICch) when membrane potential was held at -20 mV, and the mean amplitude of ICch was 102±15 pA (n = 20). 2. By the conventional whole-cell patch-clamp technique, the muscarinic current was also induced by 0.5 mmol/L GTP?S (Guanosine -5'-[?-thio] triphosphate, an activator of GTP ??in the pipette solution when membrane potential was held at -20 mV. After dialysing of GTP?S from the pipette solution for 3~5 minutes, the inward current was induced and reached a peak level at about the 8th minute under the isosmotic condition without Carbachol. The mean amplitude of GTP?S - induced current was 56.8±6.4 pA (n = 10 ). 3. Using the same technique when membrane potential was held at -20 mV, the hyposmotic solution (202 mOsm) with 50 ?mol/L Carbachol made membrane stretch and it increased ICch by 145±27% (P <0.05, n = 8).4. When membrane potential was held at -20 mV, the hyposmotic solution (202 mOsm) without Carbachol made membrane stretch and it increased GTP?S-induced current from 56.8±6.4 pA to 160.8±18.1 pA. It was increased by 183±30%(P <0.05, n = 8).5. When 20 μmmol/L cytochalasin-B (Cyt-B, a microfilament disruptor) being applied to the pipette under isosmotic condition with 50 ?mol/L Carbachol, there was no significant difference between control group and Cyt-B group of ICch (Control group: 102±15 pA; Cyt-B group: 100±7.4 pA, P >0.05, n = 8).6. In the pipette solution with 20 μmmol/L Cyt-B, the hyposmotic solution with 50 ?mol/L Carbachol made membrane stretch and it increased the ICch only by 70±6%. A significant difference between control group (145±27%) and Cyt-B (70±6%) group was found (P <0.05, n = 8 ). 7. When 20μmmol/L phalloidin (a microfilament stabilizer) being applied into the pipette and 50 ?mol/L Carbachol in isosmotic solution, the ICch decreased significantly (Control group: 102±15 pA; Phalloidin group: 74±5.4 pA, P<0.05, n = 10).8. In the pipette solution with 20 μmmol/L phalloidin, the hyposmotic solution with 50 ?mol/L Carbachol made membrane stretch and it increased ICch by 545±81%. A significant difference between control group (145±27%) and phalloidin group (545±81%) was found ( P <0.05, n = 10 ).Conclusions are as follows:1. Under isosmotic condition, both Carbachol (Cch) and GTP?S could induce muscarinic current in the gastric antra...
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